TY - JOUR T1 - Efficiency of plasmid delivery and expression after lipid-mediated gene transfer to human cells in vitro. JF - Journal of Pharmacology and Experimental Therapeutics JO - J Pharmacol Exp Ther SP - 1462 LP - 1469 VL - 279 IS - 3 AU - Z Bebök AU - A M Abai AU - J Y Dong AU - S A King AU - K L Kirk AU - G Berta AU - B W Hughes AU - A S Kraft AU - S W Burgess AU - W Shaw AU - P L Felgner AU - E J Sorscher Y1 - 1996/12/01 UR - http://jpet.aspetjournals.org/content/279/3/1462.abstract N2 - Cationic liposome-mediated gene transfer has become increasingly important in the development of experimental therapies for human diseases, such as melanoma, human immunodeficiency virus infection, cystic fibrosis and alpha-1 antitrypsin deficiency. However, very little is known about the mechanisms by which lipid-mediated gene transfer occurs. We studied the kinetics of plasmid delivery and expression by using this technique. Plasmid entry in the cystic fibrosis respiratory epithelial cell line 2CFSME0-1 as well as in two other cell lines (HeP 2g and HeLa) occurred in 95 to 100% of cells within 1 hr of the initiation of lipid-mediated gene transfer. In hepatic and respiratory cells, transcription of a construct containing the cystic fibrosis transmembrane conductance regulator was observed in more than 80% of the cell population; similarly high levels of plasmid utilization were obtained in studies of HLA-B7 expression in human melanoma cells. Studies directly relevant to current human trials of lipid-mediated gene transfer indicate that plasmid entry, transcription and translation are often surprisingly efficient, and may occur in nearly 100% of human cells in culture when sensitive methods for detection are used. Furthermore, conventional X-gal immunohistochemistry markedly underestimates transfection efficiency during transient gene expression. These studies point to a new mechanistic understanding of the features that limit expression by using cationic liposomes. ER -